The cystic fibrosis transmembrane conductance regulator (CFTR), which forms adenosine 3′,5′-monophosphate (cAMP)-regulated chloride channels, is defective in patients with cystic fibrosis. This protein contains two putative nucleotide binding domains (NBD1 and NBD2) and an R domain. CFTR in which the R domain was deleted (CFTRΔR) conducted chloride independently of the presence of cAMP. However, sites within CFTR other than those deleted also respond to cAMP, because the chloride current of CFTRΔR increased further in response to cAMP stimulation. In addition, deletion of the R domain suppressed the inactivating effect of a mutation in NBD2 (but not NBD1), a result which suggests that NBD2 interacts with the channel through the R domain.